We have previously shown that the early antibody response to HIV infection is primarily to gag proteins when measured by western blot (WB) assay. Published studies using recombinant env protein suggests that the antibody response to env may occur simultaneously to or prior to a gag response. This study was undertaken to determine the nature of these differences. Recombinant env gpl6O was electrophoresised and transfered to cellulose nitrate using standard WB buffers, air dried and cut into strips. An equivalent concentration of gp160 in PBS was spotted onto the bottom of the test strips was washed and allowed to dry. The relative concentrations of WB and spotted gpl60 was determined using colloidal gold protein stain. Strips were then blocked and used to determine the specificity of antibodies in 6 well characterized seroconversion panels using a goat anti-human IgG (L&H) coupled to alkaline phosphotase. The results showed that in 2 out of 6 seroconversion panels that nondenatured antigen was detected 7 and 5 days prior to detection with WB denatured antigen. These data suggest that in some individuals early env antibodies are directed to confirmational epitopes. Comparison with licensed WB suggested that nondenatured env responses occurred earlier or at the same time as gag responses.